1. Field of the Inventin
This invention relates to and has among its objects the provision of novel compositions for therapeutic use. Further objects of the invention will be evident from the following description wherein parts and percentages are by weight unless otherwise specified.
2. Description of the Prior Art
Fibronectin or cold insoluble globulin has been the subject of several recent reviews, editorials and clinical studies (Mossesson, et al, Blood, 56, 145-158, 1980; Saba et al, Amer. J. Med., 68, 577-594, 1980; Grause, J.A.M.A., 244, 173, 1980; Robbins et al, The Amer. Surg., 663-673, December 1980; Scoville et al, Ann. Surg., 188, 521-528, 1979). This substance, which is produced by a variety of cells including endothelial cells, epithelial cells and peritoneal macrophages is probably critical as a major component of the intracellular matrix. Circulating fibronectin is a glycoprotein molecule having a molecular weight of approximately 450,000 and is made up of two chains of approximately equal size linked by disulfide bonds (Mossesson et al, ibid.). Fibronectin is felt to be important in mediating cell to cell adhesion, cell to substrate anchoring and spreading, regulating cell locomotion, mediating attachment of fibrinogen and macrophages to collagen and acting as an opsonin or attachment factor for certain particles (above references). Severe trauma or major burns result in a decrease in plasma fibronectin and a concomitant opsonic defect which can be corrected by infusion of cryoprecipitate containing fibronectin (Robbins et al, ibid. and Scoville et al, ibid.). Such therapy has resulted in a marked improvement in cardiopulmonary function, correction of shock and normalization of other hematologic parameters suggesting an important role for fibronectin in organ and microvascular integrity (Scoville et al, ibid.).
The term reticuloendothelial system (RES) is used to refer to a system of mononuclear cells, scattered throughout the body, but with the common ability to engulf (phagocytize) blood-borne particulate matter. These cells, all derived from circulating monocytes, are found throughout many organ systems, but are especially concentrated as fixed tissue cellular elements in the spleen, lymph nodes, liver, and spleen. The RES functions as one of the body's primary host defense mechanisms for clearing and detoxifying the blood of particulate matter, including fibrin, fibrin monomers, endotoxin, platelet aggregates, damaged cellular elements, bacteria, viruses, and antibody-antigen complexes. Failure of this important host defense system leads to retention of potentially harmful particulates in the blood which may result in increased vascular permeability and edema or microvascular occlusion, both of which end in organ dysfunction.
Other functions ascribed to the RES include (1) antigen-antibody processing and interaction with lymphoid cells, a step required for normal antibody production and development of normal cellular immunity, (2) host defense against spontaneously arising tumors, (3) regulation of bone marrow hematopoiesis, (4) wound healing, and (5) remodeling of bone.
Depression of RES-mediated host defense mechanisms are seen in post-operative surgical patients, in patients who have sustained severe burns or trauma, in patients with bacterial infection, neoplasia, disseminated intravascular coagulation (DIC), and in other diseases of altered immunity. Such defects in host defense may result in sepsis and multiorgan failure. Failure in RES function is believed to result, in part, from a deficiency of plasma fibronectin which has been found to be depressed in these conditions. Fibronectin is also known to be involved in the process of wound healing.
Along with IgG, IgM, C3b and C5b fibronectin acts as an opsonin (attachment or ingestion factor) for a variety of different particles (opsonic activity). Initial assays for this opsonic activity involved the use of rat liver slices and radiolabeled gelatin coated liquid emulsion. Heparin was found to be a critical component in this system and prevention of alcohol denaturation of the fibronectin molecule by the addition of beta-mercaptoethanol was essential. Later, Molnar et al, Fed. Proc., 38, 303, 1979, using trypsin treated and untreated rat liver slices found that gelatin coated latex particles were bound to liver cells by fibronectin but were not ingested. Bevilacqua et al, J. Exp. Med., 153 42-60, 1981, subsequently found that fibronectin mediates attachment, but not ingestion, of gelatin coated red blood cells to human monocytes or peritoneal macrophages. For activity in this system, the fibronectin requires interaction with denatured collagen or gelatin and the presence of magnesium ions. Monocytes attached to a fibronectin-gelatin substrate bound four times as many red blood cells sensitized with IgG, IgM or C3b than did monocytes attached to a plastic surface. Such attached monocytes expressed an increased number of Fc and C3b receptors. Thus, while not promoting ingestion itself, fibronectin may augment Ig and C3b mediated internalization (Bevilacqua et al, ibid.).
Additional studies by Kuusela, Nature, 276, 718-720, 1978, indicate that labeled fibronectin binds strongly to Staphylococcus aureus. This effect was blocked by unlabeled fibronectin, plasma containing fibronectin, D-glucosamine, D-galactosamine, L-lysine, sodium chloride and urea. Divalent cations were not required for fibronectin binding to this organism. A subsequent report by Proctor et al, Clin. Res., 27, 650A, 1979, indicated that labeled fibronectin bound strongly to S. aureus and Micrococcus luteus but not E. coli. This was associated with enhanced S. aureus and M. luteus induced neutrophil chemiluminescence and bactericidal activity. Since these initial reports, two conflicting articles have appeared in the literature. One by Doran et al, Inf. and Immun., 33, 683-689, 1981, indicated that fibronectin probably binds to protein A on the surface of staphylococci since binding of labeled fibronectin was directly proportional to the cellular protein A content of several strains. Furthermore, binding was inhibited by the addition of soluble protein A (up to 50%). These data were in contrast, however, to those reported by Kuusela, ibid., and Verbrugh et al, Inf. and Immun., 33, 811-819, 1981, who found no association between fibronectin binding and protein A content and could not inhibit binding with up to 100 micrograms/ml of this protein. This latter group also failed to demonstrate a significant role for fibronectin in promoting actual uptake of radiolabeled staphylococci by human polymorphonuclear leukocytes (PMNs) monocytes (MNs) or alveolar macrophages in the presence or absence of human serum containing antibody. It should be pointed out, however, that these latter studies were not designed to detect an effect of fibronectin on particle attachment. Thus, there is controversy about the actual role that fibronectin has in the host defense mechanism.
Proctor et al, Blood, 59, 681-687, 1982, have shown that fibronectin mediated the attachment of S. aureus to human PMNs but it did not promote PMN phagocytosis of bacteria.